In recent years, CD1d-restricted natural killer T cells (NK T cells) have been identified as a critical population of regulatory T cells that function to control autoimmune responses. Defects in these cells have been implicated as contributing factors in the progression of autoimmune islet beta cell destruction in type 1 diabetes, both in humans and in nonobese diabetic (NOD) mice. A synthetic glycolipid known as KRN7000, which is a form of alpha-galactosyl ceramide (alphaGalCer), has the capacity to specifically activate NK T cells. Treatment of NOD mice with this compound can significantly delay or prevent diabetes. Available data indicate that KRN7000 exerts its beneficial effects by stimulating NK T cells to produce protective Th2-type cytokines such as IL-4. However, KRN7000 also triggers production of Th1-type cytokines by NK T cells and by secondary stimulation of dendritic cells and natural killer cells. This could limit its therapeutic efficacy in diseases associated with a harmful Th1 inflammatory response such as type 1 diabetes. Dr. Porcelli's laboratory has recently identified structural analogues of alphaGalCer that stimulate altered responses of NK T cells, such as the preferential secretion of Th2 cytokines without concurrent Th1 cytokines. Our central hypothesis is that analogues of alphaGalCer that have an enhanced capacity to stimulate the production of Th2 cytokines will have an even greater therapeutic efficacy in diabetes prevention than alphaGalCer, and that the beneficial effects of alphaGalCer and its analogues are due at least in part to the suppression of cytotoxic T lymphocytes (CTL) specific for islet beta-cell autoantigens. In this collaborative partnership, the Principal Investigators (Drs. Porcelli and DiLorenzo) will work together to identify glycolipid activators of NK T cells that efficiently block or reverse the autoimmune-mediated destruction of pancreatic islets in the NOD mouse model of type 1 diabetes. The mechanisms by which these glycolipids alter the repertoire and effector functions of autoreactive T cells will be analyzed, and treatment protocols that optimize the use of these potential therapeutic agents in vivo will be developed. The project will thus combine Dr. DiLorenzo's established expertise in analysis of T cell responses and disease progression in the NOD model with Dr. Porcelli's expertise in the areas of glycolipid chemistry, CD1, and NK T cell biology. The Specific Aims are: (1) To test the ability of novel analogues of alphaGalCer that elicit altered response of NK T cells to prevent or reverse diabetes in NOD mice; (2) To assess the ability of these alphaGalCer analogues to promote syngeneic islet graft survival in diabetic NOD mice; (3) To determine the effects of in vivo analogue treatment on the numbers, distribution and functions of CD1d-restricted NK T cells; (4) To assess the effects of alphaGalCer analogues on the activation, expansion and distribution of diabetogenic beta-cell cytotoxic CD8+ T cells.